Purification and properties of an extracellular invertase from Azotobacter chroococcum
1991; Elsevier BV; Volume: 13; Issue: 3 Linguagem: Inglês
10.1016/0141-0229(91)90140-6
ISSN1879-0909
AutoresMercedes G. de la Vega, Francisco Javier Cejudo, A. Paneque,
Tópico(s)Enzyme Catalysis and Immobilization
ResumoAnalysis of distribution of invertase (EC 3.2.1.26) activity in Azotobacter chroococcum ATCC 4412 and A. vinelandii UW cells revealed that A. chroococcum invertase is an exocellular protein, whereas the A. vinelandii enzyme is located inside the cell. Invertase has been purified by a simple and rapid procedure from cell-free supernatants of A. chroococcum batch cultures. The enzyme preparation appeared homogeneous on SDS-PAGE (SDS-polyacrylamide gel electrophoresis) and was identified as a glycoprotein. The exoinvertase was freed of its carbohydrate component, without loss of catalytic activity, upon treatment with β-N-acetylglucosaminidase (EC 3.2.1.30). Invertase molecular weight was estimated to be 57 kDa by analytical gel filtration and 59 KDa by SDS-PAGE. It appears, therefore, to be composed of a single subunit. The amino acid analysis of A. chroococcum invertase revealed a high proportion of acidic residues and lack of arginine and histidine. 5 mm pyridoxal-5′-phosphate or pyridoxamine nearly abolished the invertase activity. Activation energy of the enzyme for sucrose hydrolysis was 18 kcal mol−1. The specific activity of A. chroococcum invertase, expressed as turnover number, was 100 s−1.
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